Cardiac muscle contraction is dependent upon a cooperative interaction between thick and thin filament sarcomeric proteins. Tropomyosin (TM), an essential thin filament protein, interacts with actin and the troponin complex to regulate contractile activity. In human striated cardiac muscle, three TM isoforms are expressed in the sarcomere: striated alpha-TM, striated beta- TM, and striated TM30 (TPM3). Alpha-TM is the principle isoform, with 30% of the total being beta-TM; the relative expression of TM30 in the human heart is unknown. There is, however, very little information regarding specific functions of these distinct isoforms. Our long-term objective is to understand the relative importance of striated muscle TM isoforms in myofilament responsiveness during mechanical and biochemical activity of normal and diseased cardiac muscle. We have developed novel transgenic and knockout mouse systems whereby controlled regulation of TM isoforms can induce heart failure and dilated cardiomyopathy and death. In another model, ablation of the striated muscle specific exons of alpha-TM from the mouse genome results in embryonic lethality. We will test the hypothesis that modification of TM alters the cooperative process by which cardiac myofilaments are turned and off which can result in abnormal cardiac function. The Specific Aims address the following questions: (1) What is the functional impact of altered properties of TM isoforms and defined mutations on myofilament activity and on the development of cardiomyopathies?; (2) What are the changes in dynamics and level of activity of heart muscle contraction and relaxation associated with these alterations?; and (3) What is the mechanism by which Tm isoform switching affects the cooperative activation of the myofilaments? The proposed work entails a comprehensive analysis and incorporates molecular, morphological, physiological, and biochemical studies to determine the role of TM during normal cardiogenesis and in the development of heart failure The Specific Aims of this project are: (1) to ascertain the functional significance of TM isoform specific amino acids in the development of heart failure; and (2) to characterize abnormal cardiac development associated with the alpha-TM homozygous null mouse and to "rescue" this mouse. Development of transgenic mouse models and selective modification of TM amino acids provides a unique opportunity to obtain essential in vivo information for understanding the role of TM in the regulation of thin filament contraction, during both normal and diseased states.